Chromatography is a process that provides for the physical separation, quantification and identification of various analytes in a sample in a single analysis. A chromatography system comprises a number of components, including a separation column that separates the sample into its individual components as the components are passed through the column by a mobile phase. The separation column includes an inlet portion that acts as the interface for input of the sample into a separation column. The mobile phase provides a carrier fluid and driving force to move the components of the sample from the inlet portion of the column to the outlet portion, with the separation of the components dependent on their interactions with an immobilized liquid or solid material within the column (stationary phase) and the mobile phase. The system also includes a detector that detects and measures components as they exit the separation column at different times. The components of the sample are measured based on the relative timing of the detection of those components at the outlet of the separation column. The exit time of a component is defined as the retention time for that component. For chromatographic methods involving large numbers of components, a significant amount of work is required to determine the retention time of each individual component during chromatographic method development. Also significant is the amount of work needed to correlate data generated on multiple instruments performing the same analysis, and on the same instrument for different samples, even for a small number of components.
In gas chromatography (GC) the carrier fluid is a gas or a supercritical fluid which acts similarly to a gas in the system. In liquid chromatography (LC) the carrier fluid is a liquid. In both gas and liquid chromatography systems a portion of the sample is injected into a steady flow of the carrier fluid at the input portion of the column, and the components are passed through the stationary phase material in the column. At the output portion of the column the individual components exit the column separately and are measured by the detector as a function of intensity (or quantity) and as a function of the time of exit relative to the sample entering the column. The column is monitored for effluent using a detector that provides a pattern of retention times which, by calibration or comparison with known samples, can be used to identify the components of the sample chemically and quantitatively.
Additional components of a chromatography system include an injector with a mixing chamber for introducing the sample into the carrier fluid at the inlet portion, fluid controls, and a computer for processing and displaying the output of the detector. The display generally identifies the output of the column formatted according to retention times.
The various parts of a chromatographic system are not static, but include systematic changes with time and use. Such changes, for example, result from environmental or other factors, including changes that may affect certain part of the system, such as, for example, the column, the stationary phase and the detector, to name a few. These changes can also affect other parts of the chromatographic system including, for example, the mobile phase, carrier fluid and the sample.
In addition to the systematic changes, a chromatographic system can experience random changes that are not predictable. These changes, for example, may result from measurement errors, improper calibration or external unpredictable stimuli, to name a few.
It is well understood that the retention time of a particular compound as measured in a first run through a chromatographic system cannot be repeated exactly in a subsequent run. The retention times vary from run to run even under the same laboratory protocols and even where special care is given to maintaining a static and controlled environment. This poses a big challenge when trying to correlate a feature in one chromatography data set with the feature of the same compound in another chromatography dataset, such as, for example, a liquid chromatography-mass spectrometry (LC-MS) dataset.
One obvious solution to this problem is to correlate corresponding features in two different datasets when both their m/z values and retention times are close to one another. The term m/z means the ratio of charge to mass of the ion detected, where z is often unity but can be a larger integer. The m/z value is used to measure peaks resulting from the detection of components of a sample that has been run through a chromatography system. This approach is prone to errors generally, and even more so when working with complicated sample chemistry. To compensate for the deviations that are usually encountered, and to cover all possible retention time deviations, a sufficiently large tolerance window can be chosen to compensate for the deviations which allows correlations to be made even when the peaks are not near one another but nevertheless fall within the tolerance window. This method of compensation, however, can introduce mismatches and errors with respect to the datasets that are output from the chromatography system. These mismatches and errors can become even more significant as the complexity of the datasets increase. The result is that features corresponding to different compounds are mistakenly correlated due to the large tolerance window.